The ultimate goal of this proposal is to understand the role of innate immunity within the context of oncolytic viral (OV) therapy for glioblastoma multiforme (GBM). OV treatment of GBM relies on cancer-specific replication of the virus leading to tumor destruction with minimal toxicity to adjacent non-neoplastic tissue. Results from the 5 clinical trials in patients with malignant glioma have shown the relative safety of this novel treatment modality. However, evidence for significant efficacy remains to be established. Our attention in this project focuses on the natural killer (NK) cell response following viral administration. Due to their antiviral properties, NK cells represent a potential barrier to OV therapy. Alternatively, the antitumor NK response has the potential of augmenting the tumor clearing properties of OV therapy. Our previously published rat in vivo studies have demonstrated that 1) the inflammatory response mediated by activated macrophages/microglia limit OV efficacy and 2) transient immune modulation with cyclophosphamide significantly enhances GBM clearance following OV inoculation. As a result, we hypothesize that NK cells coordinate a robust inflammatory response following initial OV administration that limits viral replication, spread, and tumor lysis, thereby creating a barrier to effective OV therapy for GBM. In this project, we will determine: a) how the antiviral and antitumor properties of NK cells impact overall OV therapeutic efficacy for GBM; b) how NK cells mediate macrophage/microglia inflammatory polarization; c) the key mechanistic signals that lead to NK mediated clearance of OV-infected glioma; and d) how pharmacological agents such as cyclophosphamide and valproic acid modulate the NK response following OV treatment, leading to enhanced clearance of GBM. By elucidating the role of NK cells in orchestrating the inflammatory response following OV therapy, we are highlighting a critical therapeutic target that, when hiodulated, will potentially enhance OV efficacy for GBM. Moreover, the significance of our experiments extends beyond virotherapy into the realm of virology and immunology as we attempt to discern the key mechanistic signals leading to NK mediated clearance of viral infection.